Rapid regeneration of Phaseolus angustissimus and P. vulgaris from very young zygotic embryos

Abstract

A rapid regeneration protocol for proembryos of Phaseolus angustissimus as young as 1 day after pollination (DAP) involving pod culture for 1 week followed by embryo culture for 2 weeks and embryo germination for 1 or 2 weeks is provided. Optimization of the media was conducted with pods collected 3 DAP. The best pod culture medium was composed of basal medium [(Phillips and Collins 1979) salts with (Geerts et al. 2001) vitamins], 1000 mg l⁻¹ glutamine, 1000 mg l⁻¹ casein hydrolysate, 3% sucrose and 0.5% agar. Embryo culture medium consisted of basal medium with 500 mg l⁻¹ glutamine, 250 mg l⁻¹ casein hydrolysate, 1.9 μM ABA, 3% sucrose and 0.5% bacto-agar. Embryos developed into plantlets on germination medium containing basal medium with 0.25 μM BA, 3% sucrose and 0.7% bacto-agar. Fertile, normal plants were recovered from direct embryogenesis and from micrografted embryo-derived shoots. Embryos obtained from pods collected 3 DAP regenerated plantlets at a rate of 29.3%, while embryos from pods collected 2 DAP and 1 DAP regenerated at rates of 20.2 and 4%, respectively. A second accession of P. angustissimusregenerated at a rate of 26.2%. Using this 5-week protocol for P. vulgaris resulted in a plantlet regeneration rate of 12.5%.